Design And Bioactivity Of Novel Antimicrobial Peptides And Its Interaction With Phospholipid

Antimicrobial peptides are endogenous peptide antibiotics composing important components of the innate immune system.These peptides are mainly composed by 12-50 amino acids and share some common features, carrying net positive charge, adopting an amphipathic structure upon binding to membranes and usually consisting of roughly 50% hydrophobic amino acids, which facilitate them interacting with bacterial membrane. Antimicrobial peptides have two Striking features. First, they demonstrate a broad-spectrum antimicrobial. Second, they mainly target bacterial membrane and finally result in bacterial death. Therefore, peptides are thought to be promising candidates. However, several ultimate issues such as efficacy, mechanisms and safety should be settled before the peptides can be applied. The feasibility of their widespread usage is one of the reasons of the peptide availability. Both basic research and clinical applications need high quality peptides in a cost-effective manner. However, isolation from natural sources is a unquestioning and time-consuming process.De novo peptide design is a significant method to develop new antimicrobial agents, which has been proven by many studies.Firstly, microscale thermophoresis(MST) was first applied for antimicrobial peptide-lipid interaction, providing the information with regard to the activity of the designed peptide on the molecular level. We compared the affinity of peptide I-RW with DMPG and DPMC, which peptide had high antibacterial activity and low cytotoxicity. The results show that l-RW has higher binding affinity for negatively charged DMPG. Compared to PBS in MH medium the interaction of I-RW and DMPG liposomes is stronger. Contrastiy, in RH1640, 0.9%Na Cl interaction of I-RW and DMPC liposom weaker. We can speculate in nature systems I-RW has a good selectivity to DMPG, which has a great value. As the concentration of serum increased the interaction of I-RW and DMPC weaken, affecting its application in clinical. I-RW has a stronger selective to membrane lipid of Staphylococcus aureus, consistenting with inhibition test results. We compared the affinity.of peptide I-RW with different lipid extract, the result showed that I-RW also demonstrated higher affinity for lipid extract from Staphyloccocus aureus compared to Escherichia coli and Pseudomonas aeruginosa, which were consistent with the higher antibacterial activity against Staphyloccocus aureus than Escherichia coli and Pseudomonas aeruginosa, suggesting that the binding affinity is capable to predict the antibacterial activity to some extent. Additionally, the binding of l-RW to DMPC/DPMG was also performed in fetal bovine serum solution, which showed that the components in biological solution may have interference with the binding in salt buffer. The results proved that MST is a useful and potent tool in antimicrobial peptide-lipid interaction exploration.After extensive research we found that import of aliphatic fatty acids to antimicrobial peptides lead to an increase in their antibacterial and antifungal activities and the stability of the polypeptide is easier to be decomposed. This article taken with a fatty acid chain instead of hydrophobic amino acids to design peptides. In this study, compounds was synthesized by conjunction of fatty acid with the amino acids. By studying its antibacterial activity and hemolytic explore the impact of hydrophobic of aliphatic chain, charge, location of tryptophan for the activity of lipopeptide. The results show that as the length of the fatty chain increased the antibacterial activity and hemolytic activity enhanced. Increased charge the antibacterial activity of lipopeptide enhanced, but the hemolytic activity decreased. The biologically activity of dual fatty chain lipopeptide were very low, probably due to its characteristics were extremely similar to phospholipid and so that it can not destroy the phospholipid bilayer. In addition, by exploring the interaction of lipopeptides and the analog biofilm discuss the mechanism of novel lipopeptide.By investigating the particle size of liposomes after lipopeptides interacted with DMPC/DMPG liposomes, the disruption effect of lipopeptides on biomembrane was evaluated. The results show that biological membrane were disrupted by all lipopeptides except dual fatty chain lipopeptide. Tryptophan position affect the biological activity of the lipopeptide, when it is located Between the hydrophilic region and the hydrophobic region the biological activity of lipopeptides enhanced. By fluorescence quenching experiments we can conclude that tryptophan can effectively enter into the hydrophobic cavity of the liposome, reducing the probability of collision with acrylamide. Tryptophan position did not affect the strength of its interaction with liposomes as the blue shift of fluorescence emission peak of C14-WR3 was amount to C14-R3 W. However, in the presence of liposomes the Stern–Volmer constants of C14-WR3 was larger, indicating a more exposed tryptophan in the C14-WR3. The surface pressure-area(π-A) curves of lipopeptides C12-R4,C16-R4 D,C16-R4 with DMPC/DMPG in mixed momolayer were measured by Langmnir technology. And then anatyse the miscibility, thermodynamic stability and the mixed monolayer. To study the interaction of antimicrobial peptides with phospholipid monolayer at the molecular level.We can predicte the biological activity of antimicrobial peptides based on the affinity which obtained from the MST experiment, partly. Charge, hydrophobic and other parameters control the biological activity of antimicrobial peptides jointly. Antimicrobial peptides can exhibit good biological activity when parameters reach equilibrium.This result introduced a new method to design and screen antimicrobial peptides.